昆虫学报 ›› 2023, Vol. 66 ›› Issue (5): 619-627.doi: 10.16380/j.kcxb.2023.05.002

• 研究论文 • 上一篇    下一篇

ATP合酶亚基α在棉铃虫幼虫变态中的功能机理分析

耿梓宸, 常彦鹏, 张博, 安世恒, 赵文丽*   

  1. (河南农业大学植物保护学院, 郑州 450002)
  • 出版日期:2023-05-20 发布日期:2023-06-01

Analysis of the functional mechanism of ATP synthase subunit α in the metamorphosis of Helicoverpa armigera (Lepidoptera: Noctuidae) larvae

GENG Zi-Chen, CHANG Yan-Peng, ZHANG Bo, AN Shi-Heng, ZHAO Wen-Li*   

  1. (College of Plant Protection, Henan Agricultural University, Zhengzhou 450002, China)
  • Online:2023-05-20 Published:2023-06-01

摘要:  【目的】本研究旨在解析ATP合酶亚基α(ATP synthase subunit α, ATPs-α)在棉铃虫Helicoverpa armigera变态和发育中的功能机理。【方法】PCR扩增棉铃虫ATPs-α基因开放阅读框,并进行生物信息学分析;利用qRT-PCR检测HaATPs-α在棉铃虫5龄蜕皮期和6龄第1-5天幼虫表皮、中肠和脂肪体中及外源20-羟蜕皮酮(20-hydroxyecdysone, 20E)(0.1 mg/mL)处理后6龄幼虫表皮和脂肪体中的表达量;对棉铃虫6龄幼虫注射dsHaATPs-α,分析RNAi降低HaATPs-α的表达量对幼虫发育及变态及其体内ATP含量、海藻糖和葡萄糖含量以及海藻糖酶活性的影响。【结果】棉铃虫HaATPs-α的开放阅读框长1 677 bp,棉铃虫、草地贪夜蛾Spodoptera frugiperda、斜纹夜蛾S. litura和粉纹夜蛾Trichoplusia ni这4种鳞翅目(Lepidoptera)夜蛾科(Noctuidae)昆虫的ATPs-α氨基酸序列一致性为96.56%,且亲缘关系较近。表达模式分析结果显示,HaATPs-α在6龄第3天幼虫表皮和中肠中表达量最高,在5龄蜕皮期脂肪体中出现表达高峰;20E(0.1 mg/mL)处理较对照显著上调6龄幼虫中HaATPs-α的表达量。与对照组(注射dsGFP)相比,利用RNAi敲低HaATPs-α表达量后,幼虫发育迟缓,幼虫体重显著下降,幼虫死亡率显著升高,化蛹率和成虫羽化率显著降低,ATP含量和海藻糖酶活性均显著降低,海藻糖含量显著升高,葡萄糖含量显著降低。【结论】HaATPs-α不仅控制棉铃虫ATP的产量,同时还影响着海藻糖和葡萄糖的含量,因此,HaATPs-α在幼虫变态中发挥着重要作用。本研究结果还可为将来利用ATPs-α作为有害生物防控的新靶标提供实验证据和理论依据。

关键词: 棉铃虫, ATP合酶亚基α, ATP, 可溶性海藻糖酶, 变态

Abstract:  【Aim】This study aims to analyze the functional mechanism of ATP synthase subunit α in the metamorphosis and development of Helicoverpa armigera.【Methods】The open reading frame (ORF) of ATPS-α gene of H. armigera was amplified by PCR and analyzed by bioinformatics. qRT-PCR was employed to detect the expression levels of HaATPs-α in the cuticle, midgut and fat body of larva at the 5th instar molting stage and day-1-5 of the 6th instar, and the cuticle and fat body of the 6th instar larva after the induction of exogenous 20-hydroxyecdysone (20E)(0.1 mg/mL). The 6th instar larvae of H. armigera were injected with dsHaATPs-α, and the effects of knocking down the expression level of HaATPs-α by RNAi on the development and metamorphosis of larvae, and the ATP content, trehalose content, glucose content and trehalase activity in the larva were analyzed. 【Results】The ORF length of HaATPs-α of H. armigera is 1 677 bp. The amino acid sequence identity of ATPs-α in H. armigera, Spodoptera frugiperda, S. litura and Trichoplusia ni of Noctuidae of Lepidoptera was 96.56%, and their evolution relationship was close. The expression pattern analysis result showed that the expression levels of HaATPs-α in the cuticle and midgut of the day-3 6th instar larvae were the highest and also peaked in the fat body of the 5th instar larvae at the molting stage. Treatment with 20E(0.1 mg/mL) significantly upregulated the expression level of HaATPs-α in the 6th instar larvae compared with the control. Compared with the control group (injected with dsGFP), knocking down the expression level of HaATPs-α by RNAi delayed the larval development, significantly reduced larval weight, significantly increased larval mortality, significantly decreased pupation rate and adult emergence rate, significantly decreased ATP content and trehalase activity, significantly increased trehalose content, and significantly decreased glucose content. 【Conclusion】HaATPs-α not only controls ATP production, but also affects the contents of trehalose and glucose in H. armigera. Therefore, HaATPs-α plays an important role in larval metamorphosis. Furthermore, our results also provide experimental evidence and theoretical basis for using ATPs-α as a new target for pest control in the future.

Key words: Helicoverpa armigera, ATP synthase subunit α, ATP, soluble trehalase, metamorphosis